Nuclear substituted derivatives of 1-hydroxy-6-methoxyphenazine 5,10-dioxide

ABSTRACT

Nuclear substituted derivatives of 1-hydroxy-6-methoxyphenazine 5,10-dioxide which possess broad spectrum antimicrobial activity are disclosed.

United States Patent Leimgruber et al.

45] Oct. 24, 1972 NUCLEAR SUBSTITUTED DERIVATIVES OF l-HYDROXY-6- METHOXYPHENAZINE 5,10-DIOXIDE Inventors: Willy Leimgruber, Montclair; Manfred Weigele, North Caldwell, both of NJ.

Assignee: Hoffmann-La Roche Inc., Nutley,

Filed: June 8, 1970 Appl. No.: 44,555

US. Cl. .....260/267, 260/247.2 A, 260/2472 B,

Int. Cl. ..C07d 51/80 Field of Search ..260/267, 247.5 B, 247.2 A, 260/2472 B Primary Examiner-Alex Maze] Assistant Examiner-Anne Marie T. Tighe Attorney-Samuel L. Welt, Bernard S. Leon, Gerald S. Rosen, R. l-lain Swope, 'William M. Farley, Margaret C. Bogosian and Jon S. Saxe [57] ABSTRACT Nuclear substituted derivatives of l-hydroxy-6- methoxyphenazine 5,10-dioxide which possess broad spectrum antimicrobial activity are disclosed.

5 Claims, No Drawings NUCLEAR SUBSTITUTED DERIVATIVES OF 1- HYDROXY-6-METl-IOXYPHENAZINE l 0 DIOXIDE DESCRIPTION OF THE INVENTION wherein R, signifies bromine or the group CI-I Z,

wherein Z signifies a five or six membered heterocyclic ring containing a nitrogen atom and at most one further hetero atom consisting of oxygen; and R signifies lower alkyl or the. group W CHal'L-Rs wherein R signifies lower alkoxy, amino, mono-lower alkyl-arnino, di-lower alkylamino, or hydroxy-lower alkylamino.

As used herein the term lower alkyl either alone or in combination denotes straight or branched chain saturated hydrocarbon groups containing from one to seven carbon atoms inclusive such as methyl, ethyl, propyl, isopropyl, butyl, pentyl, hexyl and the like, with groups containing from one to four carbon atoms being preferred. The term lower alkoxy designates straight or branched chain alkoxy groups containing from one to seven carbon atoms, preferably from one to four carbon atoms, such as methoxy, ethoxy, propoxy and the like.

In a preferred aspect, when R, signifies thegroup CH Z, Z being defined as above, the heterocyclic ring. is saturated and represents a member selected from the 1 :include aromatic hydrocarbons, such as benzene,

wherein R is as described above.

The compounds of formula Ia are preferred because of their interesting level of biological activity.

Another class of compounds preferred because of their interesting biological activity and falling within the scope of the present invention are those wherein R, signifies the group CH Z, Z being defined as above, i.e., compounds of the formula OH I N\ on;

wherein R and Z are as describedv above. Most preferred of the compounds of formula I arei 6-Methoxy-2-pyrrolidinylmethyl-1 -phenazinol 5 l 0- dioxide; and V 2-Bromo-6-methoxyl -phenazinol 5 1 O-dioxide.

The compounds of formula 1 above may be prepared following a variety of procedures. The choice of i procedure will depend upon the nature of the R, and R substituents to be introduced into the phenazine j molecule.

For example, the compounds of formula I above wherein R, signifies bromine can be prepared following any conveniently available bromination technique,

; such as treating myxin or an analogue or derivative I thereof which is unsubstituted in the ortho position to the phenolic hydroxy group with bromine. For the purposes of this process aspect of the invention the bromination reaction is expediently effected in the presence of an inert organic solvent. Suitable solvents toluene and the like and halogenated hydrocarbons such as chloroform, carbontetrachloride and the like.

In a further process aspect of the present invention, the novelcompounds of formula I above wherein the R, substituent is the group CI-I Z, Z being defined as above, may be prepared by reacting myxin or an analogue or derivative thereof unsubstituted in the orthotposition to the phenolic hydroxy group with a saturated cyclic amine in the presence of formaldehyde. This reaction is expediently effected in the formamide (DMF) and di-methyl sulfoxide (DMSO),

amines that may be used in this process aspect include morpholine, piperidine, pyrrolidine and the like. Temperature and reaction time are not critical to this ?processs aspect. Thus, temperatures between about 10 C and about C are suitable, with roomtemperature being preferred. Likewise, reaction times which permit completion of the reaction are employed,

usually from about 1 to 24 hours.

1 -hydroxy-6-methoxyphenazine 5 l O-dioxide, the known antibiotic myxin, which is used as the starting material in several of the process aspects of the present invention, may be prepared following a variety of 3 procedures. One such approach involves the methylation of 1,6-phenazinediol 5,10-dioxide, the known antibiotic iodinin. This methylation procedure I is p e erab y se ies 9! n Y9 St -v In h fir ms? be utilized in the methylation step, thus eliminating the necessity of first isolating the iodinin salt. Suitable alkali metal bases that may be used to form the iodinin salt include alkali metal loweralkp xldes cgntaini ng from, one to four carbon atoms. Preferred alkali metal lower alkoxides include the sodium and potassium compounds, with potassium t-butoxide being the most preferred.

In the second stage of the methylation process, the mono alkali metal saltof iodinin is-selectively methylated by reacting said compound with conventional methylating agents. As indicated above, this selective methylation of the mono alkali metal salt of iodinin is expediently effected in the presence of an aproticpolar organic solvent such as HMPT, DMF and DMSO, with HMPT being preferred. The methylation reaction conditions can be varied. However for convenience and optimum yields suitable conditions include the use of temperatures between about C and about 70 C and reaction times sufficient to complete the reaction, usually from about i to about 24 hours. In a preferred aspect, the methylation reaction is effected at room temperature. Suitable methylating agents include methyl halides, such as methyl bromide and methyl iodide, dimethyl sulfate, and methyl tosylate.

In a further process aspect of this invention, the novel compounds of formula I wherein the R substituent is a lower alkyl group containing from 2 to 7 carbon atoms may be prepared following procedures analogous to those employed in-the preparation of myxin. For example, these compounds can be prepared by the selective alkylation of the mono alkali metal salt of iodinin. This alkylation procedure is conducted as described above for the methylation of iodinin. Suitable alkylating agents for this purpose include dialkyl sulfates, such as diethyl sulfate, lower alkyl halides, such as ethyl bromide, l-bromo butane and the like, and lower alkyl tosylates.

In another process aspect of the present invention, the novel compounds of formula I above wherein the R substituent is the group O :CIb Rt ethyl a-bromo-butyrate and the like. As indicated above, this selective alkylation of the mono alkali metal salt of iodinin is expediently effected in the presence of an aprotic polar organic solvent, such as HMPT, DMF

I and DMSO, with 'HMPT being preferred. The alkylation reaction conditions can be varied. However, again for convenience and optimum yields, suitable conditions include the use of temperatures from about 10 C to about C, with room temperature being preferred, and reaction times sufficient to complete the reaction,

' usually 1 to 24 hours.

. In a further process aspect of-the present invention the novel compounds of formula I above wherein the lg substituent is the group and R, signifies an amino group or a primary or secondary amino group may be prepared by treating the ester 'derivative, that is the corresponding compound The novel compounds of formula I have been found to possess broad spectrum antimicrobial activity. in particular, these compounds have demonstrated a high level of activity against a wide variety of bacteria, yeast and fungi such as Streptococcus agalactiae, Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, Moraxella bovis, Candida albicans, and Microsporum canis. The novel compounds of this invention are particularly useful in the treatment of animal diseases of microbial origin. When the novel compounds of formula I are employed in the treatment of microbial infec-' tions, they are conveniently utilized in combination withsuitable pharmaceutical carrier materials. These compositions are formulated by uniformly distributing the compound of formula 1 throughout a vehicle that is chemically empatible with the particular compound, non-inhibiting with respect to the active ingredients and essentially non-injurious to the body tissue under the conditions of use. When formulated into compositions suitable for topical administration, the novel compounds of this invention are preferably employed in amounts ranging from about 0.05 percent to about 1.0 percent by weight of the composition. The compounds of this invention, when employed in forms suitable for topical administration, may be utilized in varied formulations; for example, in solid formulations including finely divided powders and granular materials and liquid formulations including suspensions, concentrations, tinctures, slurries, aerosols, and the like. Further, they may be employed as creams, gels, jellies, ointments, pastes, etc.

The following examples further illustrate the scope of the invention. All temperatures given are in degrees centigrade unless indicated otherwise.

EXAMPLE 1 dioxidetmy nl 1.30 grams of potassium was I added with stirring to 2.44 grams of iodinin (0.01 m) suspended in 150 ml. of hexamethylphosphoric triamide (HMPT). The mixture was stirred at room temperature overnight. 1.4 ml. dimethyl sulfate (0.015 m) was added to the resulting emerald solution and stirring was continued. After another three hours, the reaction mixture was poured into 1,000 ml. ice water. The aqueous mixture was extracted with 3 X 500 ml. ethyl acetate. The organic extracts were washed with 2 X 300 ml water. The water washings were reextracted with 2 X 250 ml. benzene. The organic plias es were combined, dried over anhydrous magnesium sulfate, filtered and evaporated in vacuum. The residue was dissolved in 200 ml. methylene chloride. The resulting solution was filtered through a sintered glass funnel. The filtrate was applied to a chromatography column prepared from 100 grams silica gel slurried in methylene chloride. The column was developed with a mixture of methylene chloride and ethyl acetate (2:1 The fractions containing pure myxin were combined and evaporated in vacuum. The residue was redissolved in 100 ml. boiling phosphoric'triarnide (HMPT) and 3.9 grams of potassii um tertiary butyl alcoholate were placed in a flask. The reaction mixture was stirred at room temperature for 16 hours and then 6.0 ml. of ethyl bromo acetate was added. The stirring was continued at room temperature for 3 hours. The reaction mixture was poured into 2.0 liters of ice water and extracted with 3 X 500 ml. of ethyl acetate. The combined ethyl acetate extracts were washed with 500 ml. of water and filtered through a sintered glass funnel. The combined ethyl acetate extracts were dried over magnesium sulfate and concentrated in vacuo. The residue was dissolved in CH Cl ml. hot acetone. On cooling 1.781 grams of l-hydroxyfi-methoxyphenazine 5,10-dioxide crystallized giving a yield of 69 percent based on the iodinin.

EXAMPLE 2 Preparation of 6-Methoxy-2-pyrrolidinylmethyl-1- phenazinol 5 l O-dioxide 2.58 grams of myxin, ml. of hexamethyl phosphoric triamide (HMPT), 1 ml. of formalin and 1 ml. of pyrrolidine were placed in a flask and stirred at room temperature for 24 hours. The reaction mixture was diluted with 400 ml. of CHCl and extracted repeatedly with 10 per cent aqueous HCl. The combined acid extracts were washed with CHCl The acid extracts were made basic with Na CO The basic phase was then extracted with ethyl acetate. The ethyl acetate extract was washed with water dried over Na SO and evaporated. The residue was recrystallized from acetone to give the above named product as red crystals, mp 122 (dec.).

EXAMPLE 3 Preparation of 2-Bromo-6-methoxy-l-phenazinol 5 l0-dioxide To 2.58 g of myxin in 500 ml. of chloroform was added dropwise over a minute period a solution of 1.6 g. of bromine in 50 ml. of chloroform. The mixture was stirred for another 30 minutes and then filtered. The filtrate was washed with 5 percent sodium bicarbonate and water, dried over sodium sulfate, filtered and evaporated. The residue was dissolved in dichloromethane and chromatographed on silica gel. Fractions containing the desired product were collected and evaporated in vacuo. The residue was crystallized from chloroform/ether to yield pure material, mp 134 (dec.).

EXAMPLE 4 Preparation of 6-hydroxy-l-phenazinoxyacetic acid 5,10-dioxide ethyl ester 7.32 grams of iodinin, 450 ml. of hexamethyl and chromatographed over 250 grams of silica gel to yield 6-hydroxy-1-phenazinoxyacetic acid, 5,10-dioxide ethyl ester, mp 152 (dec.

EXAMPLE 5 Preparation of 7-Bromo-6-hydroxy-l-phenazinoxyacetic acid, 5,10-dioxide ethyl ester To a solution of 3.3 g. of 6-hydroxy-l-phenazinoxyacetic acidv 5,l0dioxide ethyl ester in 500 ml. of chloroform was added dropwise over a 30 minute period a solution of 1.6 g. of bromine in ml. chloroform. The mixture was stirred at room temperature for 24 hours. It was then filtered. The filtrate was washed with 5 percent aqueous NaHCO and water. After drying over magnesium sulfate it was evaporated to dryness in vacuo. The crystalline residue was slurried in ether and filtered to yield the above named product,

EXAMPLE 6 Preparation of 7-Brom0-6-hydroxy-N-(2-hydroxyethyl)- l -phenazinoxy-acetamide 5 l O-dioxide l g. of 7-bromo-6-hydroxy-l-phenazinoxyacetic acid 5,10-dioxide ethyl ester in 20 ml. of ethanolamine was stirred at room temperature for 1 hour. The reaction mixture was then diluted with chloroform, washed with dilute HCl, NaHCO and water. The chloroform solution was then dried over sodium sulfate and evaporated. The residue was slurried in ether and filtered giving the above named product, mp

We claim:

1. A compound selected from the group of compounds of the formula wherein Z signifies a fiveor six membered heterocyclic ring selected from the group consisting of morpholino, piperidino and pyrrolidino and R signifies lower alkyl or the group O CHai JRa wherein R signifies lower alkoxy, amino, monolower alkylamino, di-lower alkylamino, or hydrox- ,y -1 W ?Jl 1ia9;

7 v 2. A compound selec t eTt rom the group of compounds of the formula:

(T) OH N wherein R signifies lower alkoxy, amino, monolower alkylamino, di-lower alkylamino, or hydro);- 

2. A compound selected from the group of compounds of the formula:
 3. The compound of claim 2 wherein R3 is ethoxy, i.e., 7-bromo-6-hydroxy-1-phenazinoxyacetic acid 5,10-dioxide ethyl ester.
 4. The compound of claim 2 wherein R3 signifies hydroxy-ethylamino, i.e., 7-bromo-6-hydroxy-N-(2-hydroxyethyl)-1-phenazinoxyacetamide 5,10-dioxide.
 5. The compound of claim 1 wherein -CH2Z signifies 2-pyrrolidinylmethyl and R2 signifies methyl, i.e., 6-methoxy-2-pyrrolidnylmethyl-1-phenazinol 5,10-dioxide. 